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G A Adams

Publications and source records attributed to G A Adams.

At least 37 records · Page 2Linked to original sources
PubMed1988

A simplified approach to the analysis of subclasses of phospholipids: application to human platelets.

A procedure for the determination of the proportions of diacyl, alkenylacyl and alkylacyl subclasses of glycerophospholipids was developed. The procedure involves: (1) acid methanolysis of the phospholipid followed by Bligh/Dyer extraction of fatty acid methyl esters (FAME) derived from acyl chain types, dimethylacetals (DMA) derived from alkenyl ether chain types, and lysoalkyl phosphatidic acids (lysoalkyl-PA) derived from alkyl ether chain types; and (2) subsequent acetolysis to convert the lysoalkyl-PA to monoalkyl glycerol diacetates (MAGD). GLC analysis and quantitation (using internal standard, 21:0 FAME) of FAME, DMA and MAGD allowed calculation of the proportions of the three molecular subclasses. The methanolysis/acetolysis procedure gave an overall mean phospholipid recovery of 95 +/- 3%. Analysis of the major phospholipids in four separate preparations of fresh resting human platelets by this procedure showed the following range of molecular subclasses: phosphatidylcholine (PC), 86-92 mol % diacyl, 6-10 mol % alkylacyl and 2-3 mol % alkenylacyl; and phosphatidylethanoline (PE), 39-60 mol % diacyl, 5-8 mol % alkylacyl and 34-55 mol % alkenylacyl. The results of these subclass analyses were in general agreement with those reported in the literature.

Acetates↗
PubMed1988

Inhibition of human platelet phospholipase A2 by mono(2-ethylhexyl)phthalate.

There is evidence that the carcinogenic and teratogenic effects attributed to the plasticizer di(2-ethylhexyl)phthalate (DEHP) are due to its major metabolite mono(2-ethylhexyl)phthalate (MEHP). MEHP is also formed ex vivo by a plasma enzyme in blood products stored in polyvinyl chloride (PVC) DEHP plastic containers. People who receive large amounts of blood products, such as hemophiliacs or patients undergoing hemodialysis, cardiopulmonary bypass, or massive transfusion, are exposed to significant levels of plasticizer. In this study, the platelet was used to show that MEHP inhibits phospholipase A2 (PLA2), one of enzymes important in the release of arachidonic acid from membrane phospholipids. Arachidonate is the parent molecule for the synthesis of prostaglandins, thromboxanes, leukotrienes, and lipoxins that are made by a wide variety of cells. PLA2 was measured by the liberation of 14C-arachidonic acid from 1-stearoyl-2-[1-14C]arachidonyl-L-3-phosphatidylcholine. MEHP inhibits PLA2 activity noncompetitively in intact human platelets and lysates with a Ki of 3.7 x 10(-4) M. DEHP does not inhibit PLA2 in whole platelets. Inhibition of PLA2 by MEHP occurs at only three times the circulating level of MEHP measured in neonates undergoing exchange transfusion and 20-fold the levels experienced by patients during cardiopulmonary bypass. Therefore, infants and adult patients with multisystem failure who accumulate MEHP in their blood may be at risk for decreased platelet function.

Blood Platelets↗
PubMed1987

5-day storage of human platelet concentrates in 30 ml of plasma or artificial medium.

Optimal conditions for the storage of platelet concentrates were studied by changing 5 environmental parameters: bag composition (PL146 vs. PL732), volume of plasma (60 vs. 30 ml), anticoagulant (CPDA-1 vs. heparin), nutrient (glucose vs. fructose) and medium (plasma vs. artificial medium). A full bilevel factorial study was conducted to evaluate each variable alone and in combination with the other variables for their effects on platelet aggregation and release in response to single and pairs of stimuli. Serotonin uptake, pCO2, platelet count, lactate, glucose, pO2, pH and white blood cell concentration were also measured after 3 and 5 days of storage. Platelets that were stored in PL146 bags had reduced responses to stimulation by 3 days and markedly impaired responses after 5 days relative to platelets that were stored in PL732 bags. There was a large drop in pH and platelet responsiveness when platelets were stored in a volume of 30 ml in PL146 bags; these were not found when platelets were stored in 30 ml in PL732 bags. Replacing plasma with an artificial medium or adding fructose or heparin and calcium to plasma yielded platelets that were equally functional as routine controls in CPD-A1 plasma. It was concluded that replacement of plasma with 60 ml of artificial medium or a reduction of plasma volume with storage in PL732 bags are two possible mechanisms of obtaining more plasma from blood donations without compromising maximum platelet storage life.

Anticoagulants↗
PubMed1986

Survival and recovery of human platelets stored for five days in a non-plasma medium.

Human blood platelets were stored for five days as concentrates in 60 mL of: (a) plasma; (b) non-plasma medium with anticoagulant; and (c) non-plasma medium without anticoagulant. All preparations were equally functional when tested for platelet aggregation and release reaction in response to single agonist or synergistic pairs of agonists in vitro. Platelets stored in non-plasma medium with anti-coagulant had lower kallikrein, fibrino(gen)peptide A, lactate, and beta-thromboglobulin than did plasma controls after five days. In vivo recovery and survival of platelets stored in non-plasma medium with anticoagulant were 51.2% +/- 4.3% and 8.7 +/- 0.3 days, respectively, which were not statistically different from plasma controls of 39.2% +/- 4.9% and 7.2 +/- 0.8 days, respectively. It is concluded that platelets can be stored for five days in a non-plasma medium and still have good in vivo recoveries and survivals.

Blood Platelets↗
PubMed1985

Structural requirements of a membrane-spanning domain for protein anchoring and cell surface transport.

The membrane-spanning domain of the vesicular stomatitis virus glycoprotein (G) contains 20 uncharged and mostly hydrophobic amino acids. We created DNAs specifying G proteins with shortened transmembrane domains, by oligonucleotide-directed mutagenesis. Expression of these DNAs showed that G proteins containing 18, 16, or 14 amino acids of the original transmembrane domain assumed a transmembrane configuration and were transported to the cell surface. G proteins containing only 12 or 8 amino acids of this domain also spanned intracellular membranes, but their transport was blocked within a Golgi-like region in the cell. A G protein completely lacking the membrane-spanning domain accumulated in the endoplasmic reticulum and was secreted slowly. These experiments indicate that the size of the transmembrane domain is critical not only for membrane anchoring, but also for normal cell surface transport.

Animals↗
PubMed1985

Incorporation of a charged amino acid into the membrane-spanning domain blocks cell surface transport but not membrane anchoring of a viral glycoprotein.

The membrane-spanning domain of the vesicular stomatitis virus glycoprotein (G protein) consists of a continuous stretch of 20 uncharged and mostly hydrophobic amino acids. We examined the effects of two mutations which change the amino acid sequence in this domain. These mutations were generated by oligonucleotide-directed mutagenesis of a cDNA clone encoding the G protein, and the altered G proteins were then expressed in animal cells. Replacement of an isoleucine residue in the center of this domain with a strongly polar but uncharged amino acid (glutamine) had no effect on membrane anchoring or transport of the protein to the cell surface. Replacement of this same isoleucine residue with a charged amino acid (arginine) generated a G protein that still spanned intracellular membranes but was not transported efficiently to the cell surface. The protein accumulated in the Golgi region in about 50% of the cells, and about 20% of the cells had detectable protein levels in a punctate pattern on the cell surface. In the remaining cells the protein accumulated in a vesicular pattern throughout the cytoplasm. Models which might explain the abnormal behavior of this protein are discussed.

Amino Acid Sequence↗
PubMed1984

Platelet accumulation on collagen: drugs which inhibit arachidonic acid metabolism and affect intracellular cyclic AMP levels.

We have studied the accumulation of washed platelets on collagen-coated glass from flowing platelet-red blood cell suspensions in the presence and absence of drugs. Glass tubes were 10 cm long and the flow rate was 1 ml/min, 80 s-1. For all experiments, platelet accumulation was greatest near the tube's inlet with a continuous decrease to the exit. A common feature, of those drug treatments which lead to reduced accumulation at the inlet, was an increase in outlet accumulation when compared with outlet control values. Platelet-collagen adhesion resulted in maximal release of 3H-serotonin in the presence of agents that prevent platelet aggregation on collagen. Only drug treatment known to raise cAMP levels (PGE1 and dipyridamole) or prevent the formation of prostaglandins and thromboxanes (sulfinpyrazone, indomethacin and ASA) were found to inhibit aggregate growth. Platelet aggregation on collagen in the absence of thrombin likely proceeds through the liberation of prostaglandins and thromboxanes from surface-bound platelets into the flow where they may stimulate flow-born cells. An alternate hypothesis is that such treatments affect the release of alpha-granule components necessary for aggregation.

Alprostadil↗
PubMed1984

The presence of cysteine in the cytoplasmic domain of the vesicular stomatitis virus glycoprotein is required for palmitate addition.

The transmembrane glycoprotein (G protein) of vesicular stomatitis virus (VSV) is known to contain 1-2 mol of covalently linked fatty acid (palmitate) per mol of protein. G protein is oriented in cellular membranes such that the carboxyl-terminal 29 amino acids protrude into the cytoplasm. We have obtained expression in eukaryotic cells of mutagenized cDNA clones that encode VSV G proteins lacking portions of this cytoplasmic domain. Labeling of these truncated proteins with [3H]palmitate indicated that the palmitate might be linked to an amino acid residue within the first 14 residues on the carboxyl-terminal side of the transmembrane domain. Using oligonucleotide directed mutagenesis, we changed the single codon specifying cysteine in this domain to a codon specifying serine. Expression of this mutant gene results in synthesis of a G protein lacking palmitate. We suggest that linkage of palmitate to G protein is through the cysteine in the cytoplasmic domain and that such a linkage may occur in many viral and cellular glycoproteins. The G protein lacking palmitate is glycosylated and is transported normally to the cell surface.

Amino Acid Sequence↗
PubMed1983

Maximum fluid concentrations of materials released from platelets at a surface.

We examine the estimation of local concentrations of materials that are released from the dense and alpha-granules of platelets during accumulation of platelets upon collagen-coated glass. Platelet/red blood cell suspensions were perfused through a 1.3-mm-ID tube. Empirical data were used in a calculation procedure, based on diffusion and convection, designed to yield an upper bound on the interfacial fluid concentration (IFC) for each substance considered. The necessary empirical data are the rate of platelet accumulation and the maximum amount of material in the platelet capable of secretion. It was found that the IFC is dependent on the shear rate at the surface (G) and is proportional to G0.27. This means that an eightfold increase in flow rate would increase the IFCs approximately twofold. Serotonin, pyrophosphate, adenosine 5'-monophosphate (AMP), and adenosine 5'-triphosphate (ATP) were found not to be present in sufficient quantities to produce IFCs that could influence platelet aggregation if used alone at the IFC. A second set of materials, fibrinogen, fibronectin von Willebrand factor, and calcium, had IFCs less than their concentrations normally found in plasma. A third category, containing adenosine 5'-diphosphate (ADP) alone, had an IFC close to those known to affect platelet aggregation. The role of metabolites of arachidonic acid, which may promote or inhibit platelet aggregation, awaits further description.

Adenosine Diphosphate↗
PubMed1983

Kinetics of platelet adhesion and thrombus growth.

Epifluorescent microscopy was used to monitor the adhesion of platelets and the growth of platelet aggregates on collagen-coated glass tubes perfused with whole blood. The maximum basal length and width of the aggregate size increased linearly with time, growing symmetrically transverse to the direction of flow and asymmetrically in the plane longitudinal to the direction of flow. Aggregates had elliptical bases, with the major axis parallel to the direction of blood flow. These studies provide an experimental approach to studies of the kinetics of platelet interaction with artificial surfaces and give further support to the concept that blood flow has a major effect on the development of platelet thrombi.

Collagen↗
PubMed1981

Platelet adhesion and release: interfacial concentration of released materials.

Platelet deposition on collagen-coated glass and release from adherent platelets were studied in tube flow. Shear rates between 80 and 320 s-1 were evaluated by use of washed porcine platelets radiolabeled with 51Cr and [3H]serotonin in Tyrode solution containing albumin, apyrase, and red blood cells. Cell accumulation was highest at the tube's inlet and was directly proportional to exposure time for all positions up to 20 cm from the entrance and for times up to 800 s. Release was lowest at the inlet, 60%, and increased to 90% downstream. The accumulation and release information served as input to a calculation that yielded ADP surface concentration values at the tube's surface. These ranged from 0.6 microM at 80 s-1 to 1.8 microM at 320 s-1. In a turbidimetric device, ADP concentrations of 0.5, 1.0, and 5.0 microM were required to induce shape change, 10 and 80% aggregation, respectively. Aggregation was also obtained with subthreshold ADP and collagen in combination. We surmise that platelet shape change could have occurred adjacent to the surface in preparation for adhesion, aggregation, and release at the surface, and that ADP and collagen acted synergistically.

Adenosine Diphosphate↗